Application of representational difference analysis to genomic fragments of Marek's disease virus

Abstract

A rapid and simple method for isolation of DNA fragments of Marek's disease virus (MDV) based on representational difference analysis (RDA) was developed. Multiple viral DNA fragments, the sizes of which were restricted to 0.3 to 3.5 kbp, were simultaneously amplified after subtraction of chicken DNA from BamHI-, BglII-, EcoRI-, HindIII-, or XhoI-digested DNA fragments of MDV-infected cells. Nucleotide sequence of two RDA-derived fragments coincided with the sequence determined from direct sequencing of the MDV genome. We detected an interstrain difference in the size of restriction enzyme-digested fragments on agarose gel. This method was used on a single feather pulp to generate sufficient MDV DNA for cloning.